The invention relates to a chamber for the electrofusion of cells.
As a rule, in electrofusion, two different cells are fused together in an electric field. The term “cells” as employed within this application generally refers to biological cells, and in particular cells consisting of body tissues.
Of particular interest at the present time is, for example, the preparation of fusion products consisting of dendritic cells and tumor cells by electrofusion. Dendritic cells are immunoactive cells that may have antigens on their surfaces and that, as a function of these antigens, exhibit differing immunostimulating properties.
Fusion products from dendritic cells and tumor cells may be employed, for example, in tumor therapy. In this context, the publication by Kugler et al. in Nature Medicine, Volume 6, pages 332 to 336 (2000), describes the use of the above fusion products in the treatment of kidney carcinoma in humans.
Chambers of this type, a.k.a fusion cuvettes, in which the electrofusion of cells may be carried out, typically have a vessel for the suspension containing the cells and at least two electrodes whose electrode surfaces face each other. The electrodes are configured such that there is suspension between the electrode surfaces during operation of the chamber. When the electrodes are connected to different poles of a source of voltage, an electric field is generated between the electrode surfaces and this electric field brings about the electrofusion of the cells contained in the suspension.
The electrofusion can take place in three steps. In a first pre-alignment step, the cells contact each other by dielectrophoresis. In a second pulse step, a pulse is emitted causing the cell membranes to rupture such that cell membranes and, thus, the cells can fuse. To stabilize the fusion partners, in a third post-alignment step, the ruptured cells are kept in contact with each other by dielectrophoresis until the fusion is complete.
The aforementioned step sequence is not the only possible sequence for these steps. It is possible that only the pulse is emitted and then, if necessary a centrifugation step may follow to stabilize the fusion.
A comprehensive description of electrofusion may be found, for example, in the book by U. Zimmermann & G. Neill (“Electro manipulation of cells”, CRC Pub., Boca Raton, Fla., United States, 1996).
In electrofusion, the electric field generated in the chamber must be non-uniform. Only in a non-uniform field does the desired alignment of the cells by dielectrophoresis take place in the area of the highest field strength.
Various devices of this type are known in which non-uniform fields are generated. For example, German patent DE 3317415 describes a chamber that has a cylindrical core around which the wire-like electrodes are wound as a double helix. This chamber yields good fusion results. With such devices, however, only small volumes of suspension can be processed that may be insufficient for some applications such as, for example, the above-mentioned cancer therapy.
U.S. Pat. No. 4,578,168 discloses a chamber into which several flat wire mesh electrodes have been inserted. The disadvantage of this structure is that the cells get tangled in the mesh structures of the electrodes and are consequently difficult to remove after the fusion has been completed.
The above-mentioned publication in Nature Medicine, Volume 6 (2000) describes the use of an electroporation cuvette whose electrodes have been coated with a dielectric wax to create the desired non-uniform field. The disadvantage is that such an arrangement does not allow standardization or reproducibility.